Epstein-Barr virus (EBV) is an orally transmitted human herpesvirus that is carried by more than ninety percent of the population worldwide and is associated with both lymphoid and epithelial malignancies. The role that it plays in tumorigenesis is complex, but individuals who are immunosuppressed, particularly those infected with the human immunodeficiency virus, are at increased risk of developing EBV-associated malignancies. Part of this increased risk may reflect a loss of control of virus replication and an increase in virus load. The long term goal of the research in the laboratory is to understand how virus spreads and amplifies between and within hosts, moving between the B cells and epithelial cells which are its major targets. Although EBV can bind well to an epithelial cell that lacks CR2 and fusion of an epithelial cell mediated by virus glycoproteins gB and gHgL does not involve CR2, infection of an epithelial cell is much more efficient if CR2 is expressed. Our hypothesis is that this reflects downstream effects on intracellular transport of virus. These are much less well understood than those that occur in a B cell, but can involve interactions between the virus glycoprotein gp350 and the cell protein complement receptor type 2 (CR2 or CD21) and between the virus glycoprotein complex gHgL and the BMRF2 protein and cellular integrins. The immediate goals of the research in this application are to elucidate some of the early events in infection of an epithelial cell and test the hypothesis that virus binding to CR2 triggers important downstream events. These goals will be addressed with three specific aims. Aim one will determine the efficiency with which virus is transported to the nucleus and explore the effects of inhibitors of actin and microtubules. This will be done using a combination of real-time quantitative PCR (RT-QPCR), fluorescence in situ hybridization (FISH) and confocal microscopy. Aim two will determine the role that CR2 plays in increasing efficiency of infection of epithelial cells. This will be done by comparing virus transport to the nucleus in CR2-negative epithelial cells, CR2-positive epithelial cells and cells engineered to express CR2 that lacks a cytoplasmic tail and can no longer interact with FHOS, a formin homolog. Aim three will explore the effects of virus attachment on intracellular signaling. This will be done by analysis of phosphorylation using antibody array analysis. The effects of virus and a soluble form gp350 will be determined. EBV replication is ongoing in the oropharynx of most individuals and may lead to development of epithelial cancers. Understanding how EBV gets into the epithelial cell nucleus and the events that it triggers along the way is important for design of strategies to interrupt or ameliorate the disease that the virus can cause.